The broad goals for this proposal are to identify subunit-specific N- methyl-D-aspartate (NMDA) receptor interacting proteins, and then to characterize the functional consequences of identified interactions. This project has the potential to further elucidate specific signaling events that may be involved in the regulation of synaptic plasticity, and may describe the identification of novel molecular events involved in NMDA receptor regulation and anchoring. Interacting proteins will initially be identified through the use of the yeast two-hybrid system to screen rat brain cDNA libraries utilizing various NMDA receptor cytoplasmic carboxy-termini as "bait" constructs. Following fundamental characterization of interactions utilizing heterologous cell systems and biochemical assays, the consequences of interaction on localization will be assayed in actual neuronal systems. This will be accomplished by recombinant adenovirus mediated infection of primary cultured cortical neurons. Analysis will include characterizing altered localization patterns of endogenous receptors in the presence of overexpressed interacting proteins and selected regions of mutated NMDA receptors that may exert dominant-negative effects on localization. Additionally, the localization pattern of introduced NMDA receptors that have mutated interaction domains will also be studied.